Protecting the heart against ischemia or ischemia/reperfusion injury is one of the top health care objectives. Currently, one of the most effective mechanisms is ischemia preconditioning, which is achieved by intermittent occlusion of the coronary artery via balloon inflation. Unfortunately, though, its clinical applicability is hampered by the fact that its effectiveness is short-lived and, thus, requires prior knowledge of the time of an ischemic episode. Moreover, it is an invasive technique that carries the risk of atherosclerotic emboli. Thus, identification of factors that can mimic this process would be a great benefit to ischemic heart disease. MicroRNAs (miRNA) are posttranscriptional regulators that play a fundamental role in a cell's fate. While most of the studies in the field have focused on miRNA that are involved in cardiac organogenesis and pathogenesis, we have turned our attention to miRNA that are involved in protection against ischemic injury. We have recently identified miR-199a as an obligatory direct regulator of hypoxia-inducible factor 1 alpha (Hif- 1a) and Sirt1, two key molecules that are required for preconditioning of cells against hypoxia, and longevity, respectively. Hypoxia induces downregulation of miR-199a, which is required for upregulation of Hif-1a and Sirt1. By knocking down miR-199a in cultured myocytes or in a transgenic mouse model we were able to recapitulate the effects of hypoxia preconditioning and protect the cells against hypoxic damage. Modulating the expression of miRNA in vivo for therapeutic advantages remains a challenge. In phase I of this study we will experiment with different forms, doses, duration, and methods of delivery, of antimiR-199a in mice to establish its efficacy and specificity in reducing the levels of endogenous miRNA-199a and upregulation of its targets. These experiments are a prelude to phase II that will involve testing its effectiveness in protection of the heart against ischemic damage. The overall objective is to evaluate the potential of antimiR-199a delivery to the heart as a, preventive, preconditioning measure for the ischemic heart and, thus, establish the scientific merit and feasibility of the proposed R&D. Our hypotheses are, 1) systemic delivery of antimiR-199a to the heart will induce downregulation of endogenous miR-199a that will result in upregulation of Hif-1a and Sirt1 in all organs involved, including the heart, 2) the different modifications of the injected antimiR-199a oligo will impact its effectiveness, dose, and duration required for knockdown of endogenous miR-199a in the heart, and 3) locally delivered antimiR-199a will effectively knockdown miR-199a and induce upregulation of its targets in the injected myocardium and beyond, but not in any other organs. Our aim is to test different forms, doses, duration, and methods of delivery, of antimiR-199a in mice to establish its efficacy and specificity in reducing the levels of endogenous miRNA-199a and upregulation of its targets. PUBLIC HEALTH RELEVANCE: Protecting the heart against ischemia or ischemia/reperfusion injury is one of the top health care objectives. Currently, one of the most effective mechanisms is ischemia preconditioning, which is achieved by intermittent occlusion of the coronary artery via balloon inflation. Unfortunately, though, its clinical applicability is hampered by the fact that its effectiveness is short-lived and, thus, requires prior knowledge of the time of an ischemic episode. Moreover, it is an invasive technique that carries the risk of atherosclerotic emboli. Thus, identification of factors that can mimic this process would be a great benefit to ischemic heart disease. We have recently identified miR-199a that has the potential to fulfill that purpose, which will be investigated in this proposal.